Method for preparing metered liquid sample dilutions

ABSTRACT

Disclosed is a diluting method and apparatus which pumps a metered volume of a diluent into a conduit system, one portion of which includes a metered capillary tube bypass, into which is separately drawn concentrate. Once the system portions are filled with diluent and concentrate, a metered pumping of diluent forces diluent through the capillary bypass and forces the concentrate to mix with the diluent. The volume of mixed dilution thereby forced from the system equals the volume of the metered diluent. The structural arrangement includes valves which coact to obviate the need for between dilution rinsing of the system.

137-13, 09. 396499218 SF? '8 I v v r gi/e 1 Umted States Patent [15]3,649,218

Pontigny 1 Mar. 14, 1972 [54] METHOD FOR PREPARING METERED 3,085,7174/1963 Anscherlik ..23/270.5 X LIQUID SAMPLE DILUTIONS 3,152,117 10/1964Eijsberg ..23/271 X 3,166,096 l/1965 Lang ..23/272.8 Inventor: JatlquesPolmgny, Mommmency, 3,199,956 8/1965 Ferrari .....23/267 C France3,334,018 8/1967 Smythe... ..23/230 B 3 367 849 2/1968 Blaedel ..23/230B E [73] Asslgnee flaleah Fla 3,486,859 12/1969 Gremer ..23/230 B [22]Filed: May 7, 1969 FOREIGN PATENTS OR APPLICATIONS [21] Appl. No.:822,488

89,931 1957 Denmark ..23/272.7 807,852 1937 France ..23/267 C [30]Foreign Application Priority Data 1 Primary Examiner-Norman Yudkoff May8, 1968 France ..150945 Assistant Examiner s J Emery 521 11s. Cl...23/309, 231267 c, 23/272.7, Cass 23/258.5, 23/253 A, 23/259 A Tl T[51] Int. Cl. ..B01d 11/04, BOld 59/36 [57] BS C [58] Field of Search..23/230 A, 230 B, 230 M, 253 A, D s osed is a diluting method andapparatus which pumps a 23/267 C, 267 E, 271, 272.8, 272.7, 272.6, 309,metered volume of a diluent into a conduit system, one por- 312 A, 2585tion of which includes a metered capillary tube bypass, into which isseparately drawn concentrate. Once the system por- 5 References Citedtions are filled with diluent and concentrate, a metered pumping ofdiluent forces diluent through the capillary bypass and UNITED STATESPATENTS forces the concentrate to mix with the diluent. The volume ofmixed dilution thereby forced from the system equals the Toronto volumef the metered diluentstructural arrangement 6 322 cludes valves whichcoact to obviate the need for between d'l t' 'ns'n fth te 2,984,2505/1961 Foster ...23/272.8 In on n i g o e sys m 3,052,525 9/ 1962Vogelmann ..23/272.8 5 Claims, 1 Drawing Figure 7; a M 1; j

PAIENTEUMAR 14 I972 Invemor JACQUES A. PONTIGNY ATTYS.

METHOD FOR PREPARING METERED LIQUID SAMPLE DILUTIONS BACKGROUND OF THEINVENTION This invention is directed to method and apparatus forpreparing precise, high ratio dilutions of fluids, and is especiallywell adapted for biological processing of micro quantities of aconcentrate or sample in the absence of intersample contamination andintermediate system rinsings.

Diluting operations are common to laboratories, scientific, industrialand medical. When dilutions are prepared manually, their precisiondepends on the skill of the operator in charge of this tedious and longwork. Consequently, the use of automatic diluting apparatus has beencontemplated and employed for sometime. Those known up to the presenttime employ complicated and costly mechanisms, such as suction orpressure pumps, a complicated valve system and other delicate means.These apparatus are therefore fragile and poorly adapted to massdiluting operations, as for example, in the case of hematology in whichthe counting of white corpuscles requires a first dilution followed by asecond dilution for counting the red corpuscles with respect to a verylarge number of specimens.

Additionally, prior diluting apparatus and methods have the drawback ofdrawing inthe liquid to be diluted and discharging the diluted mixturethrough the same passage. In order to avoid pollution of a specimen bytraces of the preceding mixture, it was necessary to carry outintermediate, often long, rinsings and to provide the necessarystructure and control apparatus for this purpose, which furthercomplicated the apparatus.

Of specific interest to the present invention is the preparation ofdilutions for particle counting and analysis, primarily by apparatusknown and sold throughout the world under the trademark Coulter Counter"and disclosed, for example, in

SUMMARY OF THE INVENTION The invention remedies the prior art drawbacksand its object is to provide an automatic, simple, reliable and precisediluting method and apparatus which is easily and rapidly employed andavoids the aforementioned pollution.

The diluting apparatus according to the invention provides a firsttubular element having an inlet for the diluent and an outlet for thediluted liquid, a pumping device located at said inlet for circulating ametered volume of diluent in the first element, a second tubular elementfor the sample to be diluted including a capillary zone of givencapacity, means for filling the zone with sample, conduitsinterconnecting the first and second tubular elements at the ends of thezone, so that the zone forms a fluid bypass and means for preventing thediluent from entering the zone when the sample is introduced therein.

Further advantages and features of the invention will appear from theensuing description.

BRIEF DESCRIPTION OF THE DRAWING The single FIGURE is a diagrammaticside elevation view, partly in section, of one embodiment of a dilutingapparatus according to the invention.

DESCRIPTION OF A PREFERRED EMBODIMENT A first tubular element 1, throughwhich the diluent and dilution flows, and a second tubular element 2through which flows the sample or concentrate to be diluted, for exampleblood, comprise the two major flow portions of the apparatus. Both ofthese elements can be of a rigid material such as glass.

Located at an inlet 3 of the tubular element 1 is a metering pumpcomprising a cylinder 4, a piston 5 retained in its lower position by areturn spring 6, a cam 7, and a motor 8, for driving the cam to urge thepiston 5 toward the top of the cylinder. This pump is coupled to asource of diluent (not shown) by way of a conduit 9 having a verticalsection in which a ball valve 10 is placed. Thus, upon each cycle of thepump, a given amount of diluent is fed into the tubular element 1.

Intermediate the ends of the tubular element 1 is a vertical branch 11,at the ends of which are two capillary conduits 12 and 13. Between thetwo conduits 12 and 13, the branch 11 has a region of reduced diameteror a constriction 14. A ball valve 15 is disposed in the branch 11between the two conduits12 and 13.

Two other ball valves 16 and 17 are respectively located in the element1 upstream and downstream of the branch 11. The element 1 terminates inan outlet 18 which can empty into a dilution receiving vessel 19.

The tubular element 2 has a capillary zone 20, of given capacity, theends of which are connected to the two conduits 12 and 13. The element 2is fed with sample, such as blood, from a source 21 located at one ofits ends 22. An aspirator or equivalent device 23 is provided at theother end 24. The second tubular element 2 also includes shutofi' valves25 near its ends.

The apparatus operates in the following manner.

Initially, the tubular element 1 and the conduits 12, 13 are filled withdiluent and the tubular element 2 with concentrate sample. The valves 25are closed and the piston 5 is in its lower position.

Then the cam 7 urges the piston 5 to its upper position and thisdelivers a given amount of diluent to the element 1. This amount ofdiluent displaces an equal amount of diluent already contained in theelement 1 and the latter amount flows partly through the branch 11 and,owing to the constriction 14, partly through the bypass consisting ofthe conduit 13, the zone 20 and the conduit 12.

Since the zone 20 is capillary, the small amount of sample it initiallycontained is completely urged along as soon as the diluent starts toarrive in this zone 20. The mixture thus formed of a given amount ofdiluent and the amount of sample contained in the zone 20 drops into thevessel 19. The dilution is thus effected. To achieve a 1 to 500 redblood count dilution for use by a Coulter Counter, the capillary zone 20could have a 40 microliter volume, and the cylinder 4 would then have a20.04 milliliter volume; such that each precise dilution would have thevolume of 20.04 milliliters, of which 40 microliters is the bloodsample.

Thereafter, the cam 7 withdraws, the piston 5 is returned to its lowerposition by the spring 6, and diluent enters the cylinder 4. The valves25 are then opened and blood sample is again drawn from the source 21into the tubular element 2. The blood sample fills this element 2, andin particular the zone 20, the excess blood from the previous drawingoff being pushed along by the supply of new blood and discharged towaste by way of the aspirator 23.

When the zone 20 is filled with blood, the valves 25 are again closedand the starting position is obtained.

From the foregoing, it will be seen that only the tubular element 2comes in contact with and receives the sequence of concentrated sampleswith each sample purging the former from the capillary zone 20. Also,the tubular element 1, upstream of the ball valve 15, only comes intocontact with the diluent. Additionally, because of the constriction 14and the relatively small capacity of the zone 20 compared to the volumeof diluent, the initial influx of diluent will flush out the capillaryzone 20 past the ball valve 17, so that by the time that the entiremetered volume of diluent has moved past that point, the entire tubularelement 1, all the way to the outlet 18 will have been flushed clean ofany residue of the blood sample. Hence, interdilution and intersamplecontamination has been prevented without the need for any rinse cycles.

It should be apparent to those skilled in the art that synchronism ofthe pump, valve means 25 and the aspirator 23 can be achieved to causethe method and apparatus to be fully automatic.

Additionally, the volume of the cylinder 4 can, be adjustable, dependingupon the lowest position of the piston 5. Likewise, the volume of thecapillary zone 20 can be varied by making the zone an interchangeableelement; whereby, substitution of different zone elements of differentvolume can be accomplished.

What is claimed and sought to be protected by United States LettersPatent is:

l. A method for sequentially preparing precise uncontaminated liquiddilutions from liquid sample concentrate, each sequence comprising thesteps of:

filling a first conduit with diluent,

filling a second capillary conduit with liquid sample, a metered portionof the second conduit forming a liquid bypass of the first conduit,

forcing a metered volume of liquid diluent into and through the firstconduit, diverting a portion of said liquid diluent from the firstconduit and causing it to pass through the metered portion of saidsecond conduit and carry therewith a metered volume of liquid sampleinto the flowing diluent in the first conduit,

obtaining from the first conduit a volume of dilution equal to themetered volume of liquid diluent and including substantially all of theliquid sample in the metered portion of the second conduit, and

flushing the metered portion of said second conduit with liquid diluentto clean the same of any residue of said liquid sample.

2. A method according to claim 1 in which aspirating is employed in saidstep of filling the second conduit.

3. A method according to claim 1 in which cyclic pumping is employed insaid step of forcing the metered volume of diluent.

4. A method according to claim 1 in which said first conduit comprises atubular element.

5. A method according to claim 4 in which constricting the first tubularelement just downstream of the entrance to the bypass is employed insaid step of diverting and causing a portion of the liquid diluent topass through the bypass.

2. A method according to claim 1 in which aspirating is employed in saidstep of filling the second conduit.
 3. A method according to claim 1 inwhich cyclic pumping is employed in said step of forcing the meteredvolume of diluent.
 4. A method according to claim 1 in which said firstconduit comprises a tubular element.
 5. A method according to claim 4 inwhich constricting the first tubular element just downstream of theentrance to the bypass is employed in said step of diverting and causinga portion of the liquid diluent to pass through the bypass.